红毛五加多糖对实验性肝损伤的保护作用

红毛五加多糖能明显降低小鼠和大鼠四氯化碳、硫代乙酰胺及D-氨基半乳糖肝损伤所致血清谷丙转氨酶的升高,使磺溴酞钠滞留量减少;尚能使正常和四氯化碳中毒小鼠戊巴比妥钠诱导的睡眠时间显著缩短。     

Bone Sialoprotein Promotes Tumor Cell Migration in Both In Vitro and In Vivo Models

The present study was conducted to determine the effects of bone sialoprotein (BSP) in promoting vascular invasion of tumor cells in metastasis. We used a Matrigel system and the MDA-231 human breast cancer cells transfected with human BSP cDNA (MDA-231/BSP). Quantative analysis indicated an average of 1.7-fold increase in cell numbers that migrated through the endothelial cells in MDA-231/BSP cells compared with empty vector-transfected MDA-231 cells (MDA-231/EV). In an in vivo assay, the MDA-231 cells were incubated with or without BSP antibodies and were then inoculated onto the upper chorioallantoic membrane (CAM) of chicken embryos, in which the only route for the tumor cells to reach the lower CAM was to migrate through the embryonic vasculature. PCR amplification using human Alu primers and genomic DNA from harvested lower CAM showed an average reduction of 67% in the samples treated with BSP antibodies. These preliminary data suggest that, in metastasis, BSP may enhance the penetrating ability of tumor cells through endothelial cells and basement membrane into blood vessels. BSP antibodies can specifically hinder this effect in an in vivo system.     

Correlation between calcification and bone sialoprotein and osteopontin in papillary thyroid carcinoma

The correlation between calcification and papillary thyroid carcinoma has received increasing attention. We investigated the ability of bone sialoprotein (BSP) and osteopontin (OPN) protein levels to diagnose papillary thyroid carcinoma (PTC), and explored the correlation between BSP and OPN protein levels and calcification in PTC. Archival PTC specimens from patients with PTC with calcification and lateral cervical lymph node metastasis (LNM) were included in this retrospective immunohistochemical study. The protein levels of BSP and OPN were analysed immunohistochemically using routinely prepared tissue sections. PTC specimens from 66 patients with PTC were reviewed retrospectively (25 patients with histological calcification seen in paraffin sections, 41 patients without calcification; 35 patients with lateral cervical LNM, 31 patients without LNM). The percentage of samples that had cells that demonstrated positive protein staining differed significantly between PTC specimens, benign thyroid nodules, and adjacent normal follicular epithelium (BSP: 87.88%, 55.00%, and 42.50%, respectively; OPN: 83.33%, 70.00% and 50.00%, respectively). There was a significant difference in the immunohistochemical score (IHS) for BSP and OPN protein staining between PTC specimens with and without calcification (P < 0.05). The level of BSP protein staining was found to be significantly correlated with the level of OPN protein staining in PTC specimens. We conclude that the strong correlation between BSP and OPN and PTC suggests a role for BSP and OPN in calcification and tumor progression of PTC. BSP and OPN might be useful tumour markers for the diagnosis of PTC with limited value, because both of them had low specificity.     

Down-regulation of MeCP2 in Hirschsprung's disease

Background/Purpose

Hirschsprung's disease (HSCR) is a congenital disorder characterized by the absence of intramural ganglion cells which are highly associated with impaired proliferation and migration of neural crest cells. Whether methyl CpG binding protein 2 (MeCP2) is related with HSCR still remains unknown. This study investigates the involvement of MeCP2 in HSCR.

Methods

Quantitative real time PCR and western blot were used to detect the expression level of MeCP2 both in the aganglionic/diseased segment and the ganglionic/normal segment. In vitro assays we used siRNAs to knock-down the expression of MeCP2 in SH-SY5Y cell lines, and furthermore, MTT and transwell assays were used to detect the proliferation and migration ability, respectively. In addition, bisulfite sequencing (BSP) and miRNA analysis were used to examine why MeCP2 is decreased in HSCR samples.

Results

MeCP2 exhibited a lower expression level in tissues of HSCR patients compared with the controls. The down-regulation may also suppress the proliferative ability of the cells. However, there was no significant difference in the MeCP2 methylation level between cases and controls. Similarly, there was no difference between cases and controls in miRNA-34b (miR-34b) which is predicted to regulate MeCP2 through complementary binding to the 3′-untranslated region of MeCP2.

Conclusion

Our results indicated that an aberrant decreased level of MeCP2 may play an important role in the pathogenesis of HSCR.     

Studies on the mechanism of paracetamol-induced protection against paracetamol hepatotoxicity.

In rats, 3 days treatment with paracetamol (1 oral dose of 1 g/kg daily) produced a complete protection against the hepatotoxic actions of a further dose of paracetamol as documented by determination of serum enzyme activities (glutamic-oxaloacetic transaminase, (GOT), glutamic-pyruvic transaminase (GPT), sorbitol dehydrogenase (SDH), bromsulphthalein retention and histological investigations. Subacute paracetamol treatment decreased liver glutathione levels by 46%, liver microsomal cytochrome P-450 content by 23%, hepatic hydroxylation of aniline by 29% and hepatic demethylation of aminopyrine by 46%. It afforded also some protection against the hepatotoxic actions of carbon tetrachloride, bromobenzene and thioacetamide, but did not influence the antiphlogistic activity of paracetamol (carrageenan paw edema test). Plasma and liver concentrations of free paracetamol after oral administration of 1 g/kg paracetamol were somewhat higher in the subacutely paracetamol-pretreated rats than in the non-pretreated control animals whereas no differences in the concentrations of conjugated paracetamol were found between the 2 groups. Pretreatment with paracetamol did not influence the urinary excretion of free paracetamol but caused some shift in the urinary excretion of paracetamol conjugates: pretreated rats excreted 23% less of the paracetamol glucuronide and sulfate and 33% more of the paracetamol mercapturate than the control animals. A depression of the microsomal mixed-function oxidase activity is presumed to be the main cause of the paracetamol-induced protection against paracetamol hepatotoxicity.     

Preventive effect of silymarin against taurolithocholate-induced cholestasis in the rat

Increased amounts of monohydroxylated bile salts (BS) have been found in neonatal cholestasis, parenteral nutrition-induced cholestasis and Byler's disease, among others. We analyzed whether the hepatoprotector silymarin (SIL), administered i.p. at the dose of 100mg/kg/day for 5 days, prevents the cholestatic effect induced by a single injection of the model monohydroxylated BS taurolithocholate (TLC, 30 micromol/kg, i.v.) in male Wistar rats. TLC, administered alone, reduced bile flow, total BS output, and biliary output of glutathione and HCO(3)(-) during the peak of cholestasis (-75, -67, -81, and -80%, respectively, P<0.05). SIL prevented partially these alterations, so that the drops of these parameters induced by TLC were of only -41, -25, -60, and -64%, respectively (P<0.05 vs. TLC alone); these differences between control and SIL-treated animals were maintained throughout the whole (120 min) experimental period. Pharmacokinetic studies showed that TLC decreased the intrinsic fractional constant rate for the canalicular transport of both sulfobromophthalein and the radioactive BS [14C]taurocholate by 60 and 68%, respectively (P<0.05), and these decreases were fully and partially prevented by SIL, respectively. SIL increased the hepatic capability to clear out exogenously administered TLC by improving its own biliary excretion (+104%, P<0.01), and by accelerating the formation of its non-cholestatic metabolite, tauromurideoxycholate (+70%, P<0.05). We conclude that SIL counteracts TLC-induced cholestasis by preventing the impairment in both the BS-dependent and -independent fractions of the bile flow. The possible mechanism/s involved in this beneficial effect will be discussed.     

白及多糖对矽肺大鼠模型治疗作用的实验研究

目的：观察白及多糖的抗矽肺作用。方法：采用暴露式气管注入法,在大鼠两气管软骨环之间,注入50mg／mL的无菌石英粉混悬液1mL制备大鼠矽肺模型。将实验大鼠随机分成5组,即假手术组、模型组及白及多糖高、中、低3个剂量组。白及多糖高、中、低剂量组大鼠灌胃给予白及多糖400、200、及100mg／kg,连续10w,假手术组、模型组灌胃给予等量生理盐水。结果：白及多糖能显著降低矽肺大鼠的肺湿重,能显著降低肺组织中羟脯氨酸的含量。结论：白及多糖可在一定程度上延缓或抑制矽肺病变的发展,其作用机制与抑制肺组织中羟脯氨酸的生成有关。     

Recombinant expression and affinity purification of a novel epididymal human sperm-binding protein, BSPH1

Mammalian sperm undergo a series of maturation steps beforeacquiring fertilization competence. Our previous work demonstratedthe importance of binder of sperm (BSP) proteins in bovine spermcapacitation. Recent studies identified a BSP-homologous DNAsequence in the human genome (BSPH1) and mRNA expression inthe epididymis. The aim of this study was to develop an efficientmethod to express and purify recombinant human BSPH1. BSPH1accumulates in inclusion bodies when expressed with an N-terminalhexahistidine tag in BL21 (DE3) Escherichia coli cells. Similarto other BSP proteins, BSPH1 contains two fibronectin type-II(Fn2) domains, each consisting of two disulfide bonds. Therefore,when expressed in Origami B (DE3)pLysS cells, a strain favouringdisulfide bond formation, an improvement in soluble proteinyield was observed. However, protein was aggregated, which complicatedsubsequent purification steps. Expression of glutathione-S-transferase-taggedBSPH1 in both cell types also led to accumulation in inclusionbodies. Finally, successful production of soluble and activeprotein was achieved when BSPH1 was expressed as a His₆-thioredoxin-taggedprotein. Recombinant protein bound phosphatidylcholine liposomes,low-density lipoproteins and human sperm, therefore displayedbinding activities common to all BSP-family proteins, whichmay indicate similar biological function(s). This approach wasalso successful in producing the murine orthologue of BSPH1in the soluble and active form. Thus, fusion to thioredoxinand expression in Origami B (DE3)pLysS cells may constitutea strategy applicable to all BSP-family proteins, and possiblyto other proteins containing Fn2 domains. This work is importantto elucidate the role of BSPH1 in human sperm functions andfertility.     

Biochemical markers in oncology. Part I: molecular basis. Part II: clinical uses

The investigation of the molecular mechanisms involved in carcinogenesis and tumor progression has led to the development of numerous biochemical markers. Biochemical markers may serve for early prediction of tumor recurrence, progression and development of metastases including bone metastases and for prediction of response to therapy. Tumor antigens have been used for more than a decade and although they have shown promising clinical results, their sensitivity and specificity remain limited. A lot of knowledge on the key molecules which control cell cycle, apoptosis and angiogenesis has been acquired during recent years, but their clinical value remains uncertain. Molecular markers which are linked to malignant transformation may provide a non-surgical therapeutic approach by targeting these molecules through gene therapy or antisense molecules. Because of the complexity of the physiopathogical processes involved in tumorogenesis and metastases, we first provide a review on the molecular basis of the various tumor markers and then discuss their potential clinical utility for the major cancers. The review of the current literature indicates that at the exception of a few examples, such as the use of Her-2 to predict response of the targeted Herceptin therapy, no single marker is sensitive and specific enough to perform an accurate diagnosis, predict disease progression or response to treatment. A combination of different biochemical and imaging markers appears to be the most promising strategy to monitor patients with cancer.     

Properties of flavonoids influencing the binding to bilitranslocase investigated by neural network modelling

Bilitranslocase is a plasma membrane carrier firstly identified on the sinusoidal (vascular) domain of liver cells and later on also in the gastric epithelium. It transports diverse organic anions, such as bilirubin, some phthaleins and many dietary anthocyanins, suggesting that it could play a role both in the absorption of flavonoids from dietary sources and in their hepatic metabolism. This work was aimed at characterising the interaction of bilitranslocase with flavonols, a flavonoid sub-class. The results obtained show that, contrary to anthocyanins, flavonol glycosides do not interact with the carrier, whereas just some of the corresponding aglycones act as relatively poor ligands to bilitranslocase. These data point to a clear-cut discrimination between anthocyanins and flavonols occurring at the level of the bilitranslocase transport site. A quantitative structure-activity relationship based on counter propagation artificial neural network modelling was undertaken in order to shed light on the nature of flavonoid interaction with bilitranslocase. It was found that binding relies on the ability to establish hydrogen bonds, ruling out the involvement of charge interactions. This requisite might be at the basis of the discrimination between anthocyanins and flavonols by bilitranslocase and could lie behind some aspects of the distinct pharmacokinetic properties of anthocyanins and flavonols in mammals.